The invention relates to lipophile derivatives of muramylpeptides having properties of activating macrophages, and more particularly, tumoricidal properties. It relates more particularly to compositions containing these lipophile dervatives of muramylpeptides, more particularly compositions based on liposomes or which can form liposomes with which are incorporated said muramylpeptide lipophile derivatives.
The activation of macrophages is one of the principal mechanisms of antitumoral activity of immunomodulators: activated macrophages are capable of destroying syngenic tumoral cells not only in vitro, but also in vivo.
It has also been shown that muramylpeptides could increase the antitumoral activity of macrophages in vitro. It is not the same in vivo when the substances are used in saline solution. This inactivity is doubtless due to the fact that muramylpeptides penetrate slowly into the macrophages, no doubt by fluid pinocytosis and that muramylpeptides are rapidly eliminated from the organism through the kidney. It is then a consequence of these two phenomena that muramylpeptides injected in saline solution do not reach a sufficient concentration in the macrophages to be able to activate them.
Several solutions have been contemplated to overcome these drawbacks. In particular, it has been shown that encapsulation of muramylpeptides in liposomes confers on macrophages a cytotoxic activity already considerable in vitro and in vivo. FIDLER and his collaboraters have developed this approach by the use of multilamellar liposomes composed of phosphatidylcholine (PC) and phosphatidylserine (PS) in a ratio 7/3 and including MDP; thus they arrive at targeting this immunomodulator towards circulating monocytes which are differentiated, under the influence of the MDP that they have endocyted, into activated macrophages (Canc. Res., 42, 161-167 (1982)).
The composition of the liposomes and their nature (multilamellar) permit them to be directed, in particular, to the capillaries of the pulmonary circulation. The monocytes which have phagocyted them then migrate into the lung where they are differentiated in activated macrophages; these activated macrophages are capable of destroying metastases of tumors with pulmonary tropism like the B.sub.16 melanoma in the mouse.
However, the use of soluble muramylpeptides runs up against numerous drawbacks: certain liposomes (in particular those whose composition favours targeting towards the monocytes of the pulmonary circulation) "leak", that is to say they loose the encapsulated solute: this leakage is particularly marked when the PC/PS liposomes are placed in the presence of serum. These leakages naturally prevent good preservation of the liposomes.
This drawback is partly overcome by the use of multilamellar liposomes. It is possible in fact to think that the innermost interlamellar spaces can contain or limit the leakages before the phagocytosis of the liposome. This solution involves however a loss of specificity of targeting. In fact the use of unilamellar liposomes or constituted by few lamellae could present advantages on targeting at other organs than the lung (G. POSTE et Coll., Cancer Res., 42, 1412-1422 (1982).
To overcome this, FIDLER et Coll., have also already recognised the use of lipophile derivatives of N-acetyl-muramyl-L-alanyl-D-isoglutamine (MDP), or of N-acetyl-muramyl-L-alanyl-D-isoglutamyl-L-alanine (MTP), such as MTP-phosphatidylethanolamine.
It has also been proposed to conjugate MDP or MTP with phosphatidylethanolamine or with a stearic acid molecule, to stimulate the cytotoxic activity of the macrophages. However, the preparation of liposomes containing such derivatives does not proceed without difficulty. In particular, the formation of fine films of lipids containing the muramylpeptide derivatives and the transformation of these fine films of lipids into liposomes by conventional methods does not always lead to reproducible results and the liposomes can scarcely be preserved for prolonged periods.
In addition, it is observed that certain derivatives of muramylpeptides already containing certain lipophile groups, could show toxic and/or destabilising properties with respect to the macrophages.
It is an object of the invention to overcome yet more effectively the difficulties which have been mentioned above, in particular of providing novel MDP derivatives, possessing a considerable capacity of activating macrophages, and more particularly, of their in vivo tumoricidal activity, and this more particularly when they are administered in liposome form.
It is also an object of the invention to provide muramylpeptide lipophile derivatives which can be used among other things in the preparation of liposomes incorporating them, these liposomes then being characterised simultaneously by particularly high properties of stimulating macrophages and by great stability, both as regards the possibilities of storing these liposomes (or of compositions which can easily be transformed into liposomes) and their absence of destabilising or toxic action with respect to macrophages.